<p>Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer with poor prognosis. Treatment options of TNBC are limited, which is prone to recurrence and metastasis, posing a serious threat to patients' health. Recent studies revealed the important role of ferroptosis in TNBC. Vanillin is a phenolic aldehyde extracted from natural vanilla pods with multiple pharmacological activities, including anti-tumor properties. The present study aimed to explore the potential of vanillin in treating TNBC from the perspective of inducing ferroptosis. BT549 and MDA-MB-231 cells were incubated with 8 and 16 μM Vanillin for 24 h. Significantly decreased colony number, enhanced ROS, MDA, and GSSG levels, and reduced GSH levels and SOD activities in BT549 and MDA-MB-231 cells were triggered by 8 and 16 μM Vanillin, accompanied by increased Fe²⁺ levels, upregulated ACSL4 and KLF2, and downregulated GPX4. MDA-MB-231 cells were transfected with lentivirus containing si-KLF2 for 48 h or incubated with Fer-1 (an inhibitor of ferroptosis, 10 μM), followed by treated by 16 μM Vanillin for 24 h. Repressed cell viability, enhanced MDA and GSSG levels, declined GSH levels and SOD activities, increased Fe²⁺ levels, upregulated ACSL4, and downregulated GPX4 observed in Vanillin-treated MDA-MB-231 cells were significantly abolished by silencing KLF2 or Fer-1. Collectively, Vanillin suppressed proliferation of TNBC cells by inducing ferroptosis via mediating the KLF2/GPX4 axis.</p>